Project Summary/Abstract The objective of this project is to utilize high dimensional single cell approaches to delineate the spatial architecture of human bone marrow. Functions of hematopoietic stems cells are known to be modulated by signals generated by stromal cells in the bone marrow. Geographic relationships between hematopoietic cells and specific stromal cell populations are thought to be a critical component of this interaction. While studies in animal models have elucidated many of these key relationships, similar studies in human bone marrow have been limited. We therefore propose to utilize imaging mass cytometry (IMC) to directly interrogate the cellular architecture of the human bone marrow microenvironment. We will couple this approach with single cell RNA sequencing to enable the identification of novel markers of specific stromal cell populations. These findings will be leveraged to further inform the IMC approach. Within this context, we will develop a road map of normal human bone marrow, which can serve as the framework for subsequent studies of both normal and diseased hematopoietic states.